Nishikawa Hanako, Sasaki Masaru, Nishiyama Ken-Ichi
The United Graduate School of Agricultural Sciences, Iwate University, Morioka, Japan.
Department of Biological Chemistry and Food Sciences, Faculty of Agriculture, Iwate University, Morioka, Japan.
Bio Protoc. 2020 May 20;10(10):e3626. doi: 10.21769/BioProtoc.3626.
It is important to experimentally determine how membrane proteins are integrated into biomembranes to unveil the roles of the integration factors, and to understand the functions and structures of membrane proteins. We have developed a reconstitution system for membrane protein integration in using purified factors, in which the integration reaction is highly reproducible. This system enabled not only analysis of membrane-embedded factors including glycolipid MPIase, but also elucidation of the detailed mechanisms underlying membrane protein integration. Using the system, the integration of membrane proteins can be evaluated through a protease-protection assay. We report here how to prepare (proteo)liposomes and to determine the activities of membrane protein integration.
通过实验确定膜蛋白如何整合到生物膜中，以揭示整合因子的作用，并了解膜蛋白的功能和结构，这一点很重要。我们利用纯化因子开发了一种用于膜蛋白整合的重组系统，其中整合反应具有高度可重复性。该系统不仅能够分析包括糖脂MPIase在内的膜嵌入因子，还能够阐明膜蛋白整合的详细机制。使用该系统，可以通过蛋白酶保护试验评估膜蛋白的整合情况。我们在此报告如何制备（蛋白）脂质体以及如何确定膜蛋白整合的活性。
